On practical problems in estimation of antioxidant activity of compounds by DPPH method (Problems in estimation of antioxidant activity)

2,2′-Diphenyl-1-picrylhydrazyl (DPPH) method is commonly applied for the estimation of antioxidant activity of single compounds and their mixtures. The method is based on the spectrophotometric measurement of DPPH concentration changes resulting from the DPPH reaction with an antioxidant. The amount of remaining DPPH in the examined system is a measure of the antioxidative activity of compounds. The study shows that the type and amount of solvent used for the antioxidant dissolution, water content, and hydrogen or metal ion concentration in the measuring system significantly influence the differences in the amount of unreacted DPPH. The presence of ethyl acetate or dioxane or metal ions in the system decreases the kinetics of the reaction between DPPH and butylohydroxytoluene in relation to its kinetics in the system containing only methanolic solutions. The opposite effect is observed for water, low concentration of chloroform or hydrogen ions. The presented relationships confirm the necessity of standardising the DPPH method and show the complexity of the problem, even in very simple DPPH/antioxidant systems.     

Characterization and in vitro antioxidation of papain hydrolysate from black-bone silky fowl (Gallus gallus domesticus Brisson) muscle and its fractions

Black-bone silky fowl (Gallus gallus domesticus Brisson) (BSF) muscle was hydrolyzed by papain, and the hydrolysate was separated by preparative high performance liquid chromatography (HPLC). The amino acid composition of the BSF hydrolysate (BSFH) and its fractions was determined by HPLC precolumn derivation with 2,4-dinitrofluorobenzene. The molecular weight (MW) distribution of the BSFH and its fractions was measured by a peptide column on an HPLC system. Antioxidant activities of the BSFH and its fractions were studied by testing the reducing power and four radical scavenging systems: superoxide anion (O₂⁻), hydroxyl (·OH), 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS⁺) radicals. The results demonstrated that the BSFH had strong antioxidant capacity to scavenge O₂⁻, DPPH and ABTS⁺, and displayed strong reducing power, but revealed less powerful ability to scavenge ·OH. Fraction II of the BSFH exhibited the highest activity in scavenging O₂⁻ and DPPH, and reducing power, whereas fraction I displayed the strongest ·OH scavenging ability. Besides Glu, Asp and Gly, the rich amino acids of Ala and Leu played an important role in antioxidant activity. The small-size peptides with MW ranging from approximately 200-6000 Da probably contributed to higher antioxidant activity. Results from this study indicated that BSFH and its fractions could be used as food additives and diet nutrients.     

Total phenolic contents and antioxidant properties of commonly consumed vegetables grown in Colorado

A total of 38 commonly consumed vegetable samples, including 3 kale, 1 rhubarb, 3 spinach, 3 broccoli, 2 green bean, 5 carrot, 10 tomato, and 11 potato samples, were investigated for their total phenolic contents (TPC) and antioxidant properties. The measured antioxidant properties included free radical scavenging activities against DPPH, superoxide anion radical (O₂⁻), ABTS⁺ and peroxyl radical, and Fe²⁺ chelating capacity. The tested vegetables differed in their antioxidant properties and TPC, although all vegetables had significant antioxidant activities and contained significant levels of phenolics. The TPC and the measured antioxidant properties were correlated to each other. The Fe⁺² chelating capacity and the scavenging capacities against ABTS⁺, DPPH and O₂⁻ were reported for these commonly consumed vegetables for the first time. Results from this study suggest that kale, spinach, broccoli, and rhubarb are the better dietary sources of natural antioxidant activities and phenolic compounds. Furthermore, kale, broccoli and spinach produced in Colorado may have greater antioxidant contents than those grown at other locations.